Research Article
Gündüz Sefası (Ipomoea purpurea) Bitkisinden Polifenol Oksidaz Enziminin Kısmi Saflaştırılması ve Karakterizasyonu
Abstract
Bu çalışmada, polifenol oksidaz (PPO) enziminin Ipomoea purpurea (I. purpurea) bitkisinden ilk kez saflaştırılması ve enzimin biyokimyasal özelliklerinin belirlenmesi amaçlandı. Buna göre, in vitro olarak üretilen bitkiye ait yapraklardan optimum koşullarda (25 mg ml-1 Polivinilpolipirrolidon, pH 7.0) gerçekleştirilen ekstraksiyon sonrası elde edilen ham ekstrakt örneğinden PPO enzimi üçlü faz ayırma tekniği ile yaklaşık %57 aktivite geri kazanımla 10,5 kat saflaştırıldı. Enzimin optimum pH ve sıcaklık değerleri sırasıyla 7.0 ve 30°C olarak belirlendi. Aktivite boyama sonrası kısmen saf enzimde lakkaz, peroksidaz ve katekol oksidaz aktiviteleri tespit edildi. Ayrıca, 20°C ile 40°C arasında enzim aktivitesinin ≥%75, pH 7.0 ile 9.0 arasında ≥%65 oranında korunduğu tespit edildi.
Supporting Institution
Kocaeli Üniversitesi
Project Number
FMP-2021-2448
References
- Alici EH, Arabaci G 2016. Purification of Polyphenol Oxidase from Borage (Trachystemon orientalis L.) by Using Three-Phase Partitioning and Investigation of Kinetic Properties. International Journal of Biological Macromolecules 93: 1051-1056.
- Atala C, Quilodran M, Molina-Montenegro MA 2014. Induced Twining in Ipomoea purpurea (L.) Roth.: Response Threshold and Induction by Volatiles and Snail Damage. Gayana Botanica 71(2): 181-187.
- Aydemir T 2004. Partial Purification and Characterization of Polyphenol Oxidase from Artichoke (Cynara scolymus L.) Heads. Food Chemistry 87(1): 59-67.
- Bayraktar H, Önal S 2013. Concentration and Purification of α-Galactosidase from Watermelon (Citrullus vulgaris) by Three-Phase Partitioning, Separation and Purification Technology 118: 835-841.
- Bello AB, Sule MS, Alhassan AJ 2011. Optimum pH and pH Stability of Crude Polyphenol Oxidase (PPO) Extracted from Five Fruit Samples Commonly Consumed in Kano State, Nigeria. Bayero Journal of Pure and Applied Sciences 4(1): 26-31.
- Chew AK, Van Lehn RC 2019. Quantifying the Stability of the Hydronium Ion in Organic Solvents with Molecular Dynamics Simulations. Frontiers in Chemistry 7: 439.
- Dennison C, Lovrien R 1997. Three Phase Partitioning: Concentration and Purification of Proteins, Protein Expression and Purification 11(2): 149-161.
- de Oliveira Carvalho J, Orlanda JFF 2017. Heat Stability and Effect of pH on Enzyme Activity of Polyphenol Oxidase in Buriti (Mauritia flexuosa Linnaeus f.) Fruit Extract. Food Chemistry 233: 159-163.
- Derardja AE, Pretzler M, Kampatsikas I, Barkat M, Rompel A 2017. Purification and Characterization of Latent Polyphenol Oxidase from Apricot (Prunus armeniaca L.). Journal of Agricultural and Food Chemistry 65(37): 8203-8212.
- Duman Y, Kaya E 2014. Purification and Recovery of Invertase from Potato Tubers by Three Phase Partitioning and Determination of Kinetic Properties of Purified Enzyme. Turkish Journal of Biochemistry 39(4): 443-448.
- Gong Z, Li D, Liu C, Cheng A, Wang W 2015. Partial Purification and Characterization of Polyphenol Oxidase and Peroxidase from Chestnut Kernel. LWT - Food Science and Technology 60(2): 1095-1099.
- Gülçin İ, İrfan Küfrevioğlu Ö, Oktay M 2005. Purification and Characterization of Polyphenol Oxidase from Nettle (Urtica dioica L.) and Inhibitory Effects of Some Chemicals on Enzyme Activity. Journal of Enzyme Inhibition and Medicinal Chemistry 20(3): 297-302.
- Ioniţă E., Gurgu L, Aprodu I, Stănciuc N, Dalmadi I, Bahrim G, Râpeanu G 2017. Characterization, Purification, and Temperature/Pressure Stability of Polyphenol Oxidase Extracted from Plums (Prunus domestica). Process Biochemistry 56: 177-185.
- Kaptan Y 2004. Utilization of Scytalidium thermophilum Phenol Oxidase in Bioorganic Syntheses. The graduate School of Natural and Applied Sciences of Middle East Technical University, Master Thesis, 107 p.
- Kavrayan D, Aydemir T 2001. Partial Purification and Characterization of Polyphenoloxidase from Peppermint (Mentha piperita). Food Chemistry 74(2): 147-154.
- Kaya ED, Bağci O 2021. Purification and Biochemical Characterization of Polyphenol Oxidase Extracted from Kirmizi Kismis Grape (Vitis vinifera L.). Journal of Food Biochemistry 45(2): e13627.
- Kıran Acemi R, Acemi A 2019. Polymerization Degree-Dependent Changes in the Effects of in vitro Chitosan Treatments on Photosynthetic Pigment, Protein, and Dry Matter Contents of Ipomoea purpurea. The EuroBiotech Journal 3(4): 197-202.
- Kocabas DS, Ogel ZB, Bakir U 2011. Screening of Tree Leaves as Annual Renewable Green Biomass for Phenol Oxidase Production and Biochemical Characterization of Mulberry (Morus alba) Leaf Phenol Oxidases. World Journal of Microbiology and Biotechnology 27(3): 701-707.
- Kouakou TH, Kouadio YJ, Kouamé P, Waffo-Téguo P, Décendit A, Mérillon JM 2009. Purification and Biochemical Characterization of Polyphenol Oxidases from Embryogenic and Nonembryogenic Cotton (Gossypium hirsutum L.) Cells. Applied Biochemistry and Biotechnology 158(2): 285-301.
- Linsmaier EM, Skoog F 1965. Organic Growth Factor Requirements of Tobacco Tissue Cultures. Physiologia Plantarum, 18, 100-127.
- Liu L, Cao S, Yang H, Qi X 2015. Pectin Plays an Important Role on The Kinetics Properties of Polyphenol Oxidase from Honeydew Peach. Food Chemistry 168: 14-20.
- Muhammad N, Uddin N, Khan MKK, Ali N 2019. Inter Specific Genetic Diversity within The Ethnomedicinally Important Ipomoea L. species through Morphological and Biochemical Profiling. International Journal of Environmental Sciences and Natural Resources 19(3): 556012.
- Nadar S, Rathod V 2017. Ultrasound Assisted Intensification of Enzyme Activity and Its Properties: A Mini-review, World Journal of Microbiology and Biotechnology 33(9): 1-12.
- Niphadkar S, Rathod VK 2015. Ultrasoubd-Assisted Three-Phase Partitioning of Polyphenol Oxidase from Potato Peel (Solanum tuberosum). Biotechnology Progress 31(5): 1340-1347.
- Öztan D 2007. Tirosinaz Enziminin Ekstraksiyonu, Saflaştırılması ve Fenollerin Gideriminde Kullanımı. Gazi Üniversitesi Fen Bilimleri Enstitüsü, Yüksek Lisans Tezi, 126 s.
- Panadare D, Rathod VK 2018. Extraction and Purification of Polyphenol Oxidase: A Review. Biocatalysis and Agricultural Biotechnology 14: 431–437.
- Park KI, Ishikawa N, Morita Y, Choi JD, Hoshino A, Iida S 2007. A bHLH Regulatory Gene in The Common Morning Glory, Ipomoea purpurea, Controls Anthocyanin Biosynthesis in Flowers, Proanthocyanidin and Phytomelanin Pigmentation in Seeds, and Seed Trichome Formation. The Plant Journal 49(4): 641-654.
- Pelalak R, Khan A, Zare MH, Sadeghi MH, Marjani A 2021. Extraction of Ingredients from Tea Leaves Using Oxidative Enzymatic Reaction and Optimization of Extraction Conditions. Scientific reports 11(1): 1-19.
- Rachana CR, Jose V 2014. Three Phase Partitioning-a Novel Protein Purification Method. International Journal of ChemTech Research 6(7): 3467-3472.
- Rescigno A, Sanjust E, Montanar L, Sollai F, Soddu G, Rinaldi AC, Oliva S, Rinaldi A 1997. Detection of Laccase, Peroxidase and Polyphenol Oxidase on a Single Polyacrylamide Gel Electrophoresis. Analytical Letters 30: 2211-2220.
- Rocha AMCN, Morais AM 2001. Characterization of Polyphenoloxidase (PPO) Extracted from ‘Jonagored’apple. Food Control 12(2): 85-90.
- Siddiq M, Dolan KD 2017. Characterization of Polyphenol Oxidase from Blueberry (Vaccinium corymbosum L.). Food Chemistry 218: 216-220.
- Smith PK, Krohn RI, Hermanson GT, Mallia AK, Gartner FH, Provenzano MD, Fujimoto EK, Goeke NM, Olson BJ, Klenk DC 1985. Measurement of Protein Using Bicinchoninic Acid. Analytical Biochemistry 150(1): 76-85.
- Srivastava D 2017. Medicinal Plant of Genus Ipomoea: Present Scenario, Challenges and Future, Prospective. Research Journal of Recent Sciences 6(10): 23-26.
- Ünal MU 2007. Properties of Polyphenol Oxidase from Anamur Banana (Musa cavendishii). Food Chemistry 100: 909-913.
- Wang J, Jiang W, Wang B, Liu S, Gong Z, Luo Y 2007. Partial Properties of Polyphenol Oxidase in Mango (Mangifera indica L. cv. “Tainong”) Pulp. Journal of Food Biochemistry 31: 45-55.
- Yan JK, Wang YY, Qiu WY, Ma H, Wang ZB, Wu JY 2018. Three-Phase Partitioning as an Elegant and Versatile Platform Applied to Nonchromatographic Bioseparation Processes. Critical Reviews in Food Science and Nutrition 58(14): 2416-2431.
- Yuzugullu Karakus Y, Kahveci B, Acemi A, Kocak G 2020. Application of Three-Phase Partitioning to the Purification and Characterization of Polyphenol Oxidase from Antioxidant Rosemary (Rosmarinus officinalis L.). International Journal of Food Engineering 16(11): 20200118.
- Yuzugullu Karakus Y, Yildirim B, Acemi A 2021. Characterization of Polyphenol Oxidase from Fennel (Foeniculum vulgare Mill.) Seeds as a Promising Source. International Journal of Biological Macromolecules 170: 261-271.
- Zaini NA, Osman A, Hamid AA, Ebrahimpour A, Saari N 2013. Purification and Characterization of Membrane-Bound Polyphenoloxidase (mPPO) from Snake Fruit [Salacca zalacca (Gaertn.) Voss]. Food Chemistry 136(2): 407-414.
Partial Purification and Characterization of Polyphenol Oxidase Enzyme from Common-Morning Glory (Ipomoea purpurea)
Abstract
This study aimed to purify and biochemically characterize polyphenol oxidase (PPO) enzyme from the plant Ipomoea purpurea (I. purpurea) for the first time. For this purpose, the crude extract sample obtained from the extraction of in vitro cultured plant leaves under optimum conditions (25 mgmL-1 Polyvinylpolypyrrolidone, pH 7.0) was subjected to three-phase partitioning, and the PPO enzyme was 10.5-fold purified with a 57% activity recovery. The optimum pH and temperature values were determined as 7.0 and 30°C, respectively. Laccase, peroxidase, and catechol oxidase activities were observed after activity staining of partially purified enzyme. From stability tests, it was noted that more than 75% and 65% of its original activity were maintained at temperatures 20℃-40℃ and pH 7.0-9.0, respectively.
Keywords
Polyphenol oxidase Three-phase partitioning Enzyme purification Characterization Ipomoea purpurea
Project Number
FMP-2021-2448
References
- Alici EH, Arabaci G 2016. Purification of Polyphenol Oxidase from Borage (Trachystemon orientalis L.) by Using Three-Phase Partitioning and Investigation of Kinetic Properties. International Journal of Biological Macromolecules 93: 1051-1056.
- Atala C, Quilodran M, Molina-Montenegro MA 2014. Induced Twining in Ipomoea purpurea (L.) Roth.: Response Threshold and Induction by Volatiles and Snail Damage. Gayana Botanica 71(2): 181-187.
- Aydemir T 2004. Partial Purification and Characterization of Polyphenol Oxidase from Artichoke (Cynara scolymus L.) Heads. Food Chemistry 87(1): 59-67.
- Bayraktar H, Önal S 2013. Concentration and Purification of α-Galactosidase from Watermelon (Citrullus vulgaris) by Three-Phase Partitioning, Separation and Purification Technology 118: 835-841.
- Bello AB, Sule MS, Alhassan AJ 2011. Optimum pH and pH Stability of Crude Polyphenol Oxidase (PPO) Extracted from Five Fruit Samples Commonly Consumed in Kano State, Nigeria. Bayero Journal of Pure and Applied Sciences 4(1): 26-31.
- Chew AK, Van Lehn RC 2019. Quantifying the Stability of the Hydronium Ion in Organic Solvents with Molecular Dynamics Simulations. Frontiers in Chemistry 7: 439.
- Dennison C, Lovrien R 1997. Three Phase Partitioning: Concentration and Purification of Proteins, Protein Expression and Purification 11(2): 149-161.
- de Oliveira Carvalho J, Orlanda JFF 2017. Heat Stability and Effect of pH on Enzyme Activity of Polyphenol Oxidase in Buriti (Mauritia flexuosa Linnaeus f.) Fruit Extract. Food Chemistry 233: 159-163.
- Derardja AE, Pretzler M, Kampatsikas I, Barkat M, Rompel A 2017. Purification and Characterization of Latent Polyphenol Oxidase from Apricot (Prunus armeniaca L.). Journal of Agricultural and Food Chemistry 65(37): 8203-8212.
- Duman Y, Kaya E 2014. Purification and Recovery of Invertase from Potato Tubers by Three Phase Partitioning and Determination of Kinetic Properties of Purified Enzyme. Turkish Journal of Biochemistry 39(4): 443-448.
- Gong Z, Li D, Liu C, Cheng A, Wang W 2015. Partial Purification and Characterization of Polyphenol Oxidase and Peroxidase from Chestnut Kernel. LWT - Food Science and Technology 60(2): 1095-1099.
- Gülçin İ, İrfan Küfrevioğlu Ö, Oktay M 2005. Purification and Characterization of Polyphenol Oxidase from Nettle (Urtica dioica L.) and Inhibitory Effects of Some Chemicals on Enzyme Activity. Journal of Enzyme Inhibition and Medicinal Chemistry 20(3): 297-302.
- Ioniţă E., Gurgu L, Aprodu I, Stănciuc N, Dalmadi I, Bahrim G, Râpeanu G 2017. Characterization, Purification, and Temperature/Pressure Stability of Polyphenol Oxidase Extracted from Plums (Prunus domestica). Process Biochemistry 56: 177-185.
- Kaptan Y 2004. Utilization of Scytalidium thermophilum Phenol Oxidase in Bioorganic Syntheses. The graduate School of Natural and Applied Sciences of Middle East Technical University, Master Thesis, 107 p.
- Kavrayan D, Aydemir T 2001. Partial Purification and Characterization of Polyphenoloxidase from Peppermint (Mentha piperita). Food Chemistry 74(2): 147-154.
- Kaya ED, Bağci O 2021. Purification and Biochemical Characterization of Polyphenol Oxidase Extracted from Kirmizi Kismis Grape (Vitis vinifera L.). Journal of Food Biochemistry 45(2): e13627.
- Kıran Acemi R, Acemi A 2019. Polymerization Degree-Dependent Changes in the Effects of in vitro Chitosan Treatments on Photosynthetic Pigment, Protein, and Dry Matter Contents of Ipomoea purpurea. The EuroBiotech Journal 3(4): 197-202.
- Kocabas DS, Ogel ZB, Bakir U 2011. Screening of Tree Leaves as Annual Renewable Green Biomass for Phenol Oxidase Production and Biochemical Characterization of Mulberry (Morus alba) Leaf Phenol Oxidases. World Journal of Microbiology and Biotechnology 27(3): 701-707.
- Kouakou TH, Kouadio YJ, Kouamé P, Waffo-Téguo P, Décendit A, Mérillon JM 2009. Purification and Biochemical Characterization of Polyphenol Oxidases from Embryogenic and Nonembryogenic Cotton (Gossypium hirsutum L.) Cells. Applied Biochemistry and Biotechnology 158(2): 285-301.
- Linsmaier EM, Skoog F 1965. Organic Growth Factor Requirements of Tobacco Tissue Cultures. Physiologia Plantarum, 18, 100-127.
- Liu L, Cao S, Yang H, Qi X 2015. Pectin Plays an Important Role on The Kinetics Properties of Polyphenol Oxidase from Honeydew Peach. Food Chemistry 168: 14-20.
- Muhammad N, Uddin N, Khan MKK, Ali N 2019. Inter Specific Genetic Diversity within The Ethnomedicinally Important Ipomoea L. species through Morphological and Biochemical Profiling. International Journal of Environmental Sciences and Natural Resources 19(3): 556012.
- Nadar S, Rathod V 2017. Ultrasound Assisted Intensification of Enzyme Activity and Its Properties: A Mini-review, World Journal of Microbiology and Biotechnology 33(9): 1-12.
- Niphadkar S, Rathod VK 2015. Ultrasoubd-Assisted Three-Phase Partitioning of Polyphenol Oxidase from Potato Peel (Solanum tuberosum). Biotechnology Progress 31(5): 1340-1347.
- Öztan D 2007. Tirosinaz Enziminin Ekstraksiyonu, Saflaştırılması ve Fenollerin Gideriminde Kullanımı. Gazi Üniversitesi Fen Bilimleri Enstitüsü, Yüksek Lisans Tezi, 126 s.
- Panadare D, Rathod VK 2018. Extraction and Purification of Polyphenol Oxidase: A Review. Biocatalysis and Agricultural Biotechnology 14: 431–437.
- Park KI, Ishikawa N, Morita Y, Choi JD, Hoshino A, Iida S 2007. A bHLH Regulatory Gene in The Common Morning Glory, Ipomoea purpurea, Controls Anthocyanin Biosynthesis in Flowers, Proanthocyanidin and Phytomelanin Pigmentation in Seeds, and Seed Trichome Formation. The Plant Journal 49(4): 641-654.
- Pelalak R, Khan A, Zare MH, Sadeghi MH, Marjani A 2021. Extraction of Ingredients from Tea Leaves Using Oxidative Enzymatic Reaction and Optimization of Extraction Conditions. Scientific reports 11(1): 1-19.
- Rachana CR, Jose V 2014. Three Phase Partitioning-a Novel Protein Purification Method. International Journal of ChemTech Research 6(7): 3467-3472.
- Rescigno A, Sanjust E, Montanar L, Sollai F, Soddu G, Rinaldi AC, Oliva S, Rinaldi A 1997. Detection of Laccase, Peroxidase and Polyphenol Oxidase on a Single Polyacrylamide Gel Electrophoresis. Analytical Letters 30: 2211-2220.
- Rocha AMCN, Morais AM 2001. Characterization of Polyphenoloxidase (PPO) Extracted from ‘Jonagored’apple. Food Control 12(2): 85-90.
- Siddiq M, Dolan KD 2017. Characterization of Polyphenol Oxidase from Blueberry (Vaccinium corymbosum L.). Food Chemistry 218: 216-220.
- Smith PK, Krohn RI, Hermanson GT, Mallia AK, Gartner FH, Provenzano MD, Fujimoto EK, Goeke NM, Olson BJ, Klenk DC 1985. Measurement of Protein Using Bicinchoninic Acid. Analytical Biochemistry 150(1): 76-85.
- Srivastava D 2017. Medicinal Plant of Genus Ipomoea: Present Scenario, Challenges and Future, Prospective. Research Journal of Recent Sciences 6(10): 23-26.
- Ünal MU 2007. Properties of Polyphenol Oxidase from Anamur Banana (Musa cavendishii). Food Chemistry 100: 909-913.
- Wang J, Jiang W, Wang B, Liu S, Gong Z, Luo Y 2007. Partial Properties of Polyphenol Oxidase in Mango (Mangifera indica L. cv. “Tainong”) Pulp. Journal of Food Biochemistry 31: 45-55.
- Yan JK, Wang YY, Qiu WY, Ma H, Wang ZB, Wu JY 2018. Three-Phase Partitioning as an Elegant and Versatile Platform Applied to Nonchromatographic Bioseparation Processes. Critical Reviews in Food Science and Nutrition 58(14): 2416-2431.
- Yuzugullu Karakus Y, Kahveci B, Acemi A, Kocak G 2020. Application of Three-Phase Partitioning to the Purification and Characterization of Polyphenol Oxidase from Antioxidant Rosemary (Rosmarinus officinalis L.). International Journal of Food Engineering 16(11): 20200118.
- Yuzugullu Karakus Y, Yildirim B, Acemi A 2021. Characterization of Polyphenol Oxidase from Fennel (Foeniculum vulgare Mill.) Seeds as a Promising Source. International Journal of Biological Macromolecules 170: 261-271.
- Zaini NA, Osman A, Hamid AA, Ebrahimpour A, Saari N 2013. Purification and Characterization of Membrane-Bound Polyphenoloxidase (mPPO) from Snake Fruit [Salacca zalacca (Gaertn.) Voss]. Food Chemistry 136(2): 407-414.
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Details
| Primary Language | English |
|---|---|
| Subjects | Structural Biology |
| Journal Section | RESEARCH ARTICLE |
| Authors | |
| Project Number | FMP-2021-2448 |
| Publication Date | December 30, 2022 |
| Submission Date | October 11, 2021 |
| Acceptance Date | February 17, 2022 |
| Published in Issue | Year 2022Volume: 25 Issue: Ek Sayı 1 |
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